B-RAF mutant alleles associated with Langerhans cell histiocytosis, a granulomatous pediatric disease

PLoS One. 2012;7(4):e33891. doi: 10.1371/journal.pone.0033891. Epub 2012 Apr 10.


Background: Langerhans cell histiocytosis (LCH) features inflammatory granuloma characterised by the presence of CD1a+ dendritic cells or 'LCH cells'. Badalian-Very et al. recently reported the presence of a canonical (V600E)B-RAF mutation in 57% of paraffin-embedded biopsies from LCH granuloma. Here we confirm their findings and report the identification of two novel B-RAF mutations detected in LCH patients.

Methods and results: Mutations of B-RAF were observed in granuloma samples from 11 out of 16 patients using 'next generation' pyrosequencing. In 9 cases the mutation identified was (V600E)B-RAF. In 2 cases novel polymorphisms were identified. A somatic (600DLAT)B-RAF insertion mimicked the structural and functional consequences of the (V600E)B-RAF mutant. It destabilized the inactive conformation of the B-RAF kinase and resulted in increased ERK activation in 293 T cells. The (600DLAT)B-RAF and (V600E)B-RAF mutations were found enriched in DNA and mRNA from the CD1a+ fraction of granuloma. They were absent from the blood and monocytes of 58 LCH patients, with a lower threshold of sequencing sensitivity of 1%-2% relative mutation abundance. A novel germ line (T599A)B-RAF mutant allele was detected in one patient, at a relative mutation abundance close to 50% in the LCH granuloma, blood monocytes and lymphocytes. However, (T599A)B-RAF did not destabilize the inactive conformation of the B-RAF kinase, and did not induce increased ERK phosphorylation or C-RAF transactivation.

Conclusions: Our data confirmed presence of the (V600E)B-RAF mutation in LCH granuloma of some patients, and identify two novel B-RAF mutations. They indicate that (V600E)B-RAF and (600DLAT)B-RAF mutations are somatic mutants enriched in LCH CD1a(+) cells and absent from the patient blood. Further studies are needed to assess the functional consequences of the germ-line (T599A)B-RAF allele.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alleles
  • Amino Acid Sequence
  • Child
  • Child, Preschool
  • Extracellular Signal-Regulated MAP Kinases / genetics
  • Extracellular Signal-Regulated MAP Kinases / metabolism
  • Female
  • Genetic Predisposition to Disease
  • Granuloma / genetics*
  • Granuloma / metabolism
  • HEK293 Cells
  • Histiocytosis, Langerhans-Cell / enzymology*
  • Histiocytosis, Langerhans-Cell / genetics*
  • Histiocytosis, Langerhans-Cell / metabolism
  • Humans
  • Infant
  • Lymphocytes / metabolism
  • Male
  • Molecular Sequence Data
  • Monocytes / metabolism
  • Mutation*
  • Phosphorylation / genetics
  • Polymorphism, Genetic
  • Proto-Oncogene Proteins B-raf / genetics*
  • Proto-Oncogene Proteins B-raf / metabolism
  • Proto-Oncogene Proteins c-raf / genetics
  • Proto-Oncogene Proteins c-raf / metabolism
  • Sequence Homology, Amino Acid
  • Transcriptional Activation
  • Young Adult


  • BRAF protein, human
  • Proto-Oncogene Proteins B-raf
  • Proto-Oncogene Proteins c-raf
  • Extracellular Signal-Regulated MAP Kinases